1d1t
MUTANT OF HUMAN SIGMA ALCOHOL DEHYDROGENASE WITH LEUCINE AT POSITION 141
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OverviewOverview
Pyrazole and its 4-alkyl substituted derivatives are potent inhibitors for, many alcohol dehydrogenases. However, the human sigma sigma isoenzyme, exhibits a 580-fold lower affinity for 4-methylpyrazole than does the, human beta1beta1 isoenzyme, with which it shares 69% sequence identity. In, this study, structural and kinetic studies were utilized in an effort to, identify key structural features that affect the binding of, 4-methylpyrazole in human alcohol dehydrogenase isoenzymes. We have, extended the resolution of the human sigma sigma alcohol dehydrogenase, (ADH) isoenzyme to 2.5 A resolution. Comparison of this structure to the, human beta1beta1 isoenzyme structure indicated that the side-chain, position for Met141 in sigma sigma ADH might interfere with, 4-methylpyrazole binding. Mutation of Met141 in sigma sigma ADH to Leu, (sigma141L) lowers the Ki for 4-methylpyrazole from 350 to 10 microM, while having a much smaller effect on the Ki for pyrazole. Thus, the, mutagenesis results show that the residue at position 141, which lines the, substrate-binding pocket at a position close to the methyl group of, 4-methylpyrazole, directly affects the binding of the inhibitor. To rule, out nonspecific structural changes due to the mutation, the X-ray, structure of the sigma141L mutant enzyme was determined to 2.4 A, resolution. The three-dimensional structure of the mutant enzyme is, identical to the wild-type enzyme, with the exception of the residue at, position 141. Thus, the differences in 4-methylpyrazole binding between, the mutant and wild-type sigma sigma ADH isoenzymes can be completely, ascribed to the local changes in the topology of the substrate binding, site, and provides an explanation for the class-specific differences in, 4-methylpyrazole binding to the human ADH isoenzymes.
About this StructureAbout this Structure
1D1T is a Single protein structure of sequence from Homo sapiens with ZN, ACT, CAC and NAD as ligands. Active as Alcohol dehydrogenase, with EC number 1.1.1.1 Full crystallographic information is available from OCA.
ReferenceReference
Methionine-141 directly influences the binding of 4-methylpyrazole in human sigma sigma alcohol dehydrogenase., Xie PT, Hurley TD, Protein Sci. 1999 Dec;8(12):2639-44. PMID:10631979
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